Zantedeschia variety 110224-64275

ABSTRACT

A  Zantedeschia  variety designated 110224-64275 is disclosed. The invention relates to the seeds of  Zantedeschia  110224-64275, the plants of  Zantedeschia  110224-64275, plant parts of  Zantedeschia  110224-64275, and methods for producing a  Zantedeschia  plant produced by crossing  Zantedeschia  110224-64275 with itself or with another  Zantedeschia  variety. The invention also relates to methods for producing a  Zantedeschia  plant containing in its genetic material one or more genes or transgenes and the transgenic  Zantedeschia  plants and plant parts produced by those methods. This invention also relates to  Zantedeschia  varieties, or breeding varieties, and plant parts derived from  Zantedeschia  110224-64275, methods for producing other  Zantedeschia  varieties, hybrids, or plant parts derived from  Zantedeschia  110224-64275, and to the  Zantedeschia  plants, varieties, and their parts derived from use of those methods. The invention further relates to hybrid  Zantedeschia  seeds, plants, and plant parts produced by crossing the  Zantedeschia  110224-64275 with another  Zantedeschia  variety.

TECHNICAL FIELD

The present invention relates to Zantedeschia breeding, a novelZantedeschia variety designated 110224-64275 and progeny derivedtherefrom.

BACKGROUND

All publications cited in this application are herein incorporated byreference. Zantedeschia sp., are members of the Araceae family.Zantedeschia are herbaceous flowering plants that are native to Africa.The genus Zantedeschia is composed of several species such as the moreevergreen, mostly white and cool-season flowering Z. aethiopica to theAestivae colored types such as Z. albomaculata, Z. rehmannii, Z.elliotiana, Z. pentlandii, and Z. jucunda.

Zantedeschia can be propagated from seed, tubers, and tissue culture.Seed, tuber and tissue culture germination protocols for Zantedeschiaare well-known in the art.

There are numerous steps in the development of any novel, desirableplant germplasm. Plant breeding begins with the analysis and definitionof problems and weaknesses of the current germplasm, the establishmentof program goals, and the definition of specific breeding objectives.The next step is selection of germplasm that possesses the traits tomeet the program goals. The goal is to combine in a single variety animproved combination of desirable traits from the parental germplasm.These important traits may include higher seed yield, resistance todiseases and insects, better stems and roots, tolerance to drought andheat, and better commercial plant and flower quality.

Zantedeschia is an important and valuable ornamental plant. Thus, acontinuing goal of ornamental plant breeders is to develop varietieswith novel characteristics, such as color, growth habit, and hardiness.To accomplish this goal, the breeder must select and develop plants thathave traits that result in superior Zantedeschia varieties.

The foregoing examples of the related art and limitations relatedtherewith are intended to be illustrative and not exclusive. Otherlimitations of the related art will become apparent to those of skill inthe art upon a reading of the specification.

SUMMARY

The following embodiments and aspects thereof are described inconjunction with systems, tools and methods which are meant to beexemplary, not limiting in scope. In various embodiments, one or more ofthe above-described problems have been reduced or eliminated, whileother embodiments are directed to other improvements.

According to one aspect of the invention, there is provided a novelZantedeschia variety designated 110224-64275. Plants of Zantedeschia arefurther valued as breeding lines enabling the development of superiorornamental Zantedeschia plants having a range of desirable coloredspathes and superior growth performance.

In another aspect, the present invention relates to plants, seeds, andother plant parts such as pollen, ovules, embryos, protoplasts,meristematic cells, callus, pollen, leaves, ovules, anthers, cotyledons,hypocotyl, pistils, roots, root tips, spathe, spadix, seeds, petiole,rhizome, pods, berries, bulb, tuber, shoot, or stems of Zantedeschia110224-64275 or the like.

In another aspect, the present invention relates to plants ofZantedeschia 110224-64275, and to methods for producing a Zantedeschiaplant produced by crossing Zantedeschia 110224-64275 with itself oranother Zantedeschia plant, and the creation of variants by mutagenesisor transformation of Zantedeschia 110224-64275.

Thus, any such methods using Zantedeschia 110224-64275 are part of thisinvention: selfing, backcrosses, hybrid production, crosses topopulations, and the like. All plants produced using Zantedeschia110224-64275 as at least one parent are within the scope of thisinvention. Zantedeschia 110224-64275 could be used in crosses withother, different, Zantedeschia plants to produce first generation (F₁)Zantedeschia hybrid seeds and plants with superior characteristics.

In another aspect, the present invention provides for a method ofproducing a Zantedeschia plant comprising crossing a first parent plantwith a second parent plant and harvesting the resultant seed, whethereither one or both parents are Zantedeschia 110224-64275.

In another aspect, the present invention provides for single or multiplegene converted plants of Zantedeschia 110224-64275. The transferredgene(s) may preferably be a dominant or recessive allele. Preferably,the transferred gene(s) will confer such traits as herbicide tolerance,insect resistance, resistance for bacterial, fungal, or viral disease,male fertility, and male sterility. The gene may be a naturallyoccurring Zantedeschia gene or a transgene or gene introduced throughgenetic engineering techniques.

In another aspect, the present invention provides regenerable cells foruse in tissue culture of Zantedeschia variety 110224-64275. The tissueculture will preferably be capable of regenerating plants having all thephysiological and morphological characteristics of the foregoingZantedeschia plant, and of regenerating plants having substantially thesame genotype as the foregoing Zantedeschia plant. Preferably, theregenerable cells in such tissue cultures will be pollen, ovules,embryos, protoplasts, meristematic cells, callus, pollen, leaves,ovules, anthers, cotyledons, hypocotyl, pistils, roots, root tips,spathe, spadix, seeds, petiole, rhizome, pods, berries, bulb, tuber,shoot, stems or the like. Still further, the present invention providesZantedeschia plants regenerated from the tissue cultures of Zantedeschia110224-64275.

In addition to the exemplary aspects and embodiments described above,further aspects and embodiments will become apparent by study of thefollowing descriptions.

DEFINITIONS

In the description and tables herein, a number of terms are used. Inorder to provide a clear and consistent understanding of thespecification and claims, including the scope to be given such terms,the following definitions are provided:

Allele. Allele is any of one or more alternative forms for a gene, allof which related to one trait or characteristic. In a diploid cell ororganism, the two alleles of a given gene occupy corresponding loci on apair of homologous chromosomes.

Cotyledon. A cotyledon is a type of seed leaf. The cotyledon containsthe food storage tissues of the seed.

Embryo. The embryo is the small plant contained within a mature seed.

F₃. The “F₃” symbol denotes a generation resulting from the selfing ofthe F₂ generation along with selection for type and rogueing ofoff-types. The “F” number is a term commonly used in genetics, anddesignates the number of the filial generation. The “F₃” generationdenotes the offspring resulting from the selfing or self mating ofmembers of the generation having the next lower “F” number, that is, the“F₂” generation.

Gene. As used herein, “gene” refers to a segment of nucleic acid. A genecan be introduced into a genome of a species, whether from a differentspecies or from the same species, using transformation or variousbreeding methods.

Hypocotyl. A hypocotyl is the portion of an embryo or seedling betweenthe cotyledons and the root. Therefore, it can be considered atransition zone between shoot and root.

Locus. A locus confers one or more traits such as, for example, malesterility, herbicide tolerance, insect resistance, and diseaseresistance. The trait may be, for example, conferred by a naturallyoccurring gene introduced into the genome of the variety bybackcrossing, a natural or induced mutation, or a transgene introducedthrough genetic transformation techniques. A locus may comprise one ormore alleles integrated at a single chromosomal location.

Plant Diameter. Plant diameter is the spread of the plant, is measuredat the widest horizontal spread of the plant.

Plant Form. Plant form refers to the silhouette or profile of the plant,ranging from upright to semi-spreading to spreading.

Plant Height. Plant height is taken from the top of the soil to the topof the spathe and is measured in centimeters.

Progeny. As used herein, includes an F₁ Zantedeschia plant produced fromthe cross of two Zantedeschia plants where at least one plant includesZantedeschia variety 110224-64275 and progeny further includes, but isnot limited to, subsequent F₂, F₃, F₄, F₅, F₆, F₇, F₈, F₉, and F₁₀generational crosses with the recurrent parental line.

Pubescence. This refers to a covering of very fine hairs closelyarranged on the leaves, stems, and pods of the Zantedeschia plant.

Quantitative Trait Loci (QTL). Quantitative trait loci (QTL) refer togenetic loci that control to some degree numerically representabletraits that are usually continuously distributed.

RHS. RHS refers to the acronym for Royal Horticultural Society thatpublishes a color chart used in the plant industry. All RHS colorsreferred to herein are from the RHS 2007 edition.

Single Gene Converted (Conversion). Single gene converted (conversion)plants refers to plants which are developed by a plant breedingtechnique called backcrossing wherein essentially all of the desiredmorphological and physiological characteristics of a variety arerecovered in addition to the single gene transferred into the varietyvia the backcrossing technique or via genetic engineering.

Spadix. Spadix refers to a type of spike inflorescence having smallflowers borne on a fleshy stem.

Spathe. Spathe refers to the leaf-like bract surrounding the spadix.

DETAILED DESCRIPTION

Zantedeschia variety 110224-64275 is a semi-spreading plant withyellow-orange spathes and strong disease resistance to Pectobacteriumcarotovora pv. carotovora. Some of the selection criteria used forvarious generations include: plant form; leaf size, shape, color;maculation presence, shape, and density; petiole length; number of stems(peduncles); determinacy of stem development; stem length and strength;spathe color, size, and shape; spadix color and size; pollen production;seed production; tuber production; resistance to soil pathogens;post-harvest quality (stem strength, stability of spathe color andshape); overall distinctness, uniformity, and stability.

Zantedeschia 110224-64275 has shown uniformity and stability, asdescribed in the following variety description information. Zantedeschia110224-64275 has been self-pollinated a sufficient number of generationswith careful attention to uniformity of plant type and has beenincreased with continued observation for uniformity.

Zantedeschia 110224-64275 has the following morphologic and othercharacteristics (based primarily on data collected at Moss Landing,Calif.). All measurements below are given as median values.

TABLE 1 VARIETY DESCRIPTION INFORMATION Plant form: Semi-spreading Plantheight: 59.0 cm Plant diameter: 56.0 cm Leaf blade width: 15.25 cm Leafblade length: 20.25 cm Leaf blade shape: Cordate Leaf blade, color ofupper surface: RHS 139A Leaf blade, color of lower surface: RHS 137CLeaf blade, undulation of margin: Absent or very weakly expressedAttitude of leaf blade: Semi-erect Leaf blade apex: Right-angled Leafblade, maculation Present on upper surface: Leaf blade, number of Verymany maculations on upper surface: Leaf blade, shape of maculations:Round Petiole length: 32.5 cm Petiole color: RHS 137C Spathe height:11.5 cm Spathe width: 9.25 cm Spathe length: 10.5 cm Spathe color (outersurface): RHS 15A (for young) and RHS 42B (at maturity) Spathe color(inner surface) RHS 15A (for young) and RHS 42B to RHS 42C (at maturity)Spathe shape: Funnel-shaped Awl-shaped tip: Present Recurve of spathe:Moderate Spathe wrapping: Moderate Wave of spathe margin: WeakFragrance: Absent Spathe, presence of Present; very slight andindistinct; throat spot: RHS 186A to RHS 186B Spadix length: 4.0 cmSpadix width: 1.0 cm Spadix color: RHS 8A Peduncle length: 57.0 cmPeduncle color: RHS 155B Pollen amount: Abundant Pollen color: RHS 137BDisease resistance: Strong; resistant to Pectobacterium carotovora pv.carotovora

When Zantedeschia variety 110224-64275 is compared to Zantedeschiavariety ‘Fire Dancer’, Zantedeschia variety 110224-64275 has small anddiscreet leaf maculation, an indistinct/slight spathe eye, a spadixcolor of RHS 8A at maturity, and a moderate recurve of the spathe thatincreases with maturity, while ‘Fire Dancer’ has large and irregularleaf maculation, distinct spathe eyes, a spadix color of RHS 46B atmaturity, and little to no recurve of the spathe.

FURTHER EMBODIMENTS Breeding with Zantedeschia Variety 110224-64275

The complexity of inheritance influences choice of the breeding method.Backcross breeding is used to transfer one or a few favorable genes fora highly heritable trait into a desirable variety. This approach hasbeen used extensively for breeding disease-resistant varieties. Variousrecurrent selection techniques are used to improve quantitativelyinherited traits controlled by numerous genes. The use of recurrentselection in self-pollinating plants depends on the ease of pollination,the frequency of successful hybrids from each pollination and the numberof hybrid offspring from each successful cross.

Promising advanced breeding lines are thoroughly tested and compared toappropriate standards in environments representative of the commercialtarget area(s) for three or more years. The best lines are candidatesfor new commercial varieties; those still deficient in a few traits maybe used as parents to produce new populations for further selection.

These processes, which lead to the final step of marketing anddistribution, usually take from eight to twelve years from the time thefirst cross is made. Therefore, development of new varieties is atime-consuming process that requires precise forward planning, efficientuse of resources, and a minimum of changes in direction.

A most difficult task is the identification of individuals that aregenetically superior, because for most traits the true genotypic valueis masked by other confounding plant traits or environmental factors.One method of identifying a superior plant is to observe its performancerelative to other experimental plants and to a widely grown standardvariety. If a single observation is inconclusive, replicatedobservations provide a better estimate of its genetic worth.

The goal of ornamental plant breeding is to develop new, unique andsuperior ornamental varieties and hybrids. The breeder initially selectsand crosses two or more parental lines, followed by repeated selfing andselection, producing many new genetic combinations. The breeder cantheoretically generate billions of different genetic combinations viacrossing, selection, selfing and mutations. Therefore, a breeder willnever develop the same variety, or even very similar varieties, havingthe same traits from the exact same parents.

Each year, the plant breeder selects the germplasm to advance to thenext generation. This germplasm is grown under unique and differentgeographical, climatic and soil conditions and further selections arethen made during and at the end of the growing season. The varietiesthat are developed are unpredictable because the breeder's selectionoccurs in unique environments with no control at the DNA level, and withmillions of different possible genetic combinations being generated. Abreeder of ordinary skill in the art cannot predict the final resultinglines he develops, except possibly in a very gross and general fashion.The same breeder cannot produce the same variety twice by using the sameoriginal parents and the same selection techniques. Thisunpredictability results in the expenditure of large amounts of researchmonies to develop superior new Zantedeschia varieties.

Breeding programs combine desirable traits from two or more varieties orvarious broad-based sources into breeding pools from which varieties aredeveloped by selfing and selection of desired phenotypes. Pedigreebreeding is used commonly for the improvement of self-pollinatingplants. Two parents that possess favorable, complementary traits arecrossed to produce an F₁. An F₂ population is produced by selfing one orseveral F₁s. Selection of the best individuals may begin in the F₂population; then, beginning in the F₃, the best individuals in the bestfamilies are selected. Replicated testing of families can begin in theF₄ generation to improve the effectiveness of selection for traits withlow heritability. At an advanced stage of inbreeding (i.e., F₆ and F₇),the best lines or mixtures of phenotypically similar lines are testedfor potential release as new varieties.

Using Zantedeschia Variety 110224-64275 to Develop Other ZantedeschiaVarieties

Zantedeschia varieties such as Zantedeschia variety 110224-64275 aredeveloped for sales in the ornamental and cut flower market. However,Zantedeschia varieties such as Zantedeschia variety 110224-64275 alsoprovide a source of breeding material that may be used to develop newZantedeschia varieties. Plant breeding techniques known in the art andused in a Zantedeschia plant breeding program include, but are notlimited to, recurrent selection, mass selection, bulk selection,hybridization, mass selection, backcrossing, pedigree breeding,open-pollination breeding, restriction fragment length polymorphismenhanced selection, genetic marker enhanced selection, making doublehaploids, mutagenesis and transformation. Often combinations of thesetechniques are used. The development of Zantedeschia varieties in aplant breeding program requires, in general, the development andevaluation of homozygous varieties. There are many analytical methodsavailable to evaluate a new variety. The oldest and most traditionalmethod of analysis is the observation of phenotypic traits, butgenotypic analysis may also be used.

Additional Breeding Methods

One embodiment of the present invention is directed to methods forproducing a Zantedeschia plant by crossing a first parent Zantedeschiaplant with a second parent Zantedeschia plant, wherein the first orsecond Zantedeschia plant is the Zantedeschia plant from Zantedeschiavariety 110224-64275. Further, both first and second parent Zantedeschiaplants may be from Zantedeschia variety 110224-64275. Therefore, anymethods using Zantedeschia variety 110224-64275 are an embodiment ofthis invention, such as selfing, backcrosses, hybrid breeding, andcrosses to populations. Any plants produced using Zantedeschia variety110224-64275 as at least one parent are also an embodiment of theinvention. These methods are well-known in the art and some of the morecommonly used breeding methods are described herein. Descriptions ofbreeding methods can be found in one of several reference books (e.g.,Allard, “Principles of Plant Breeding” (1960); and Fehr, “BreedingMethods for Cultivar Development,” 2^(nd) ed., Wilcox editor (1987)).

The following describes breeding methods that may be used withZantedeschia variety 110224-64275 in the development of furtherZantedeschia plants. One such embodiment is a method for developing aZantedeschia 110224-64275 progeny plant in a Zantedeschia plant breedingprogram comprising: obtaining the Zantedeschia plant, or a part thereof,of Zantedeschia variety 110224-64275, utilizing said plant, or plantpart, as a source of breeding material, and selecting a Zantedeschiavariety 110224-64275 progeny plant with molecular markers in common withZantedeschia variety 110224-64275 and/or with morphological and/orphysiological characteristics selected from the characteristics listedin Table 1. Breeding steps that may be used in the Zantedeschia plantbreeding program can include for example, pedigree breeding,backcrossing, mutation breeding, and recurrent selection. In conjunctionwith these steps, techniques such as RFLP-enhanced selection, geneticmarker enhanced selection (for example, SSR markers), and the making ofdouble haploids may be utilized.

Another method involves producing a population of Zantedeschia variety110224-64275 progeny Zantedeschia plants, comprising crossingZantedeschia variety 110224-64275 with another Zantedeschia plant,thereby producing a population of Zantedeschia plants which, on average,derive 50% of their alleles from Zantedeschia variety 110224-64275. Aplant of this population may be selected and repeatedly selfed or sibbedwith a Zantedeschia variety resulting from these successive filialgenerations. One embodiment of this invention is the Zantedeschiavariety produced by this method and that has obtained at least 50% ofits alleles from Zantedeschia variety 110224-64275.

One of ordinary skill in the art of plant breeding would know how toevaluate the traits of two plant varieties to determine if there is nosignificant difference between the two traits expressed by thosevarieties. For example, see, Fehr and Walt, Principles of CultivarDevelopment, pp. 261-286 (1987). Thus, the invention includesZantedeschia variety 110224-64275 progeny Zantedeschia plants comprisinga combination of at least two Zantedeschia variety 110224-64275 traitsselected from the group consisting of those listed in Table 1 orZantedeschia variety 110224-64275 combination of traits listed in theSummary, so that said progeny Zantedeschia plant is not significantlydifferent for said traits than Zantedeschia variety 110224-64275 asdetermined at the 5% significance level when grown in the sameenvironmental conditions. Using techniques described herein, molecularmarkers may be used to identify said progeny plant as a Zantedeschiavariety 110224-64275 progeny plant. Mean trait values may be used todetermine whether trait differences are significant, and preferably thetraits are measured on plants grown under the same environmentalconditions. Once such a variety is developed, its value is substantialsince it is important to advance the germplasm base as a whole in orderto maintain or improve traits such as ornamental features, diseaseresistance, pest resistance, and plant performance in extremeenvironmental conditions.

Progeny of Zantedeschia variety 110224-64275 may also be characterizedthrough their filial relationship with Zantedeschia variety110224-64275, as for example, being within a certain number of breedingcrosses of Zantedeschia variety 110224-64275. A breeding cross is across made to introduce new genetics into the progeny, and isdistinguished from a cross, such as a self or a sib cross, made toselect among existing genetic alleles. The lower the number of breedingcrosses in the pedigree, the closer the relationship betweenZantedeschia variety 110224-64275 and its progeny. For example, progenyproduced by the methods described herein may be within 1, 2, 3, 4, or 5breeding crosses of Zantedeschia variety 110224-64275.

As used herein, the term “plant” includes plant cells, plantprotoplasts, plant cell tissue cultures from which Zantedeschia plantscan be regenerated, plant calli, plant clumps, and plant cells that areintact in plants or parts of plants, such as pollen, ovules, embryos,protoplasts, meristematic cells, callus, pollen, leaves, ovules,anthers, cotyledons, hypocotyl, pistils, roots, root tips, spathe,spadix, seeds, petiole, rhizome, pods, berries, bulb, tuber, shoot, orstems and the like.

Pedigree Breeding

Pedigree breeding starts with the crossing of two genotypes, such asZantedeschia 110224-64275 and another Zantedeschia variety having one ormore desirable characteristics that is lacking or which complementsZantedeschia variety 110224-64275. If the two original parents do notprovide all the desired characteristics, other sources can be includedin the breeding population. In the pedigree method, superior plants areselfed and selected in successive filial generations. In the succeedingfilial generations, the heterozygous condition gives way to homogeneousvarieties as a result of self-pollination and selection. Typically inthe pedigree method of breeding, five or more successive filialgenerations of selfing and selection is practiced: F₁ to F₂; F₂ to F₃;F₃ to F₄; F₄ to F₅; etc. After a sufficient amount of inbreeding,successive filial generations will serve to increase seed of thedeveloped variety. Preferably, the developed variety compriseshomozygous alleles at about 95% or more of its loci.

Backcross Breeding

Backcross breeding has been used to transfer genes for a simplyinherited, highly heritable trait into a desirable homozygous variety orinbred line which is the recurrent parent. The source of the trait to betransferred is called the donor parent. After the initial cross,individuals possessing the phenotype of the donor parent are selectedand repeatedly crossed (backcrossed) to the recurrent parent. Theresulting plant is expected to have the attributes of the recurrentparent and the desirable trait transferred from the donor parent.

In addition to being used to create a backcross conversion, backcrossingcan also be used in combination with pedigree breeding. As discussedpreviously, backcrossing can be used to transfer one or morespecifically desirable traits from one variety, the donor parent, to adeveloped variety called the recurrent parent, which has overall goodcommercial characteristics yet lacks that desirable trait or traits.However, the same procedure can be used to move the progeny toward thegenotype of the recurrent parent, but at the same time retain manycomponents of the nonrecurrent parent by stopping the backcrossing at anearly stage and proceeding with selfing and selection. For example, aZantedeschia variety may be crossed with another variety to produce afirst generation progeny plant. The first generation progeny plant maythen be backcrossed to one of its parent varieties to create a BC₁ orBC₂. Progeny are selfed and selected so that the newly developed varietyhas many of the attributes of the recurrent parent and yet several ofthe desired attributes of the nonrecurrent parent. This approachleverages the value and strengths of the recurrent parent for use in newZantedeschia varieties.

Therefore, an embodiment of this invention is a method of making abackcross conversion of Zantedeschia variety 110224-64275, comprisingthe steps of crossing a plant of Zantedeschia variety 110224-64275 witha donor plant comprising a desired trait, selecting an F₁ progeny plantcomprising the desired trait, and backcrossing the selected F₁ progenyplant to a plant of Zantedeschia variety 110224-64275. This method mayfurther comprise the step of obtaining a molecular marker profile ofZantedeschia variety 110224-64275 and using the molecular marker profileto select for a progeny plant with the desired trait and the molecularmarker profile of Zantedeschia variety 110224-64275. In one embodiment,the desired trait is a mutant gene, gene, or transgene present in thedonor parent.

Recurrent Selection and Mass Selection

Recurrent selection is a method used in a plant breeding program toimprove a population of plants. Zantedeschia variety 110224-64275 issuitable for use in a recurrent selection program. The method entailsindividual plants cross-pollinating with each other to form progeny. Theprogeny are grown and the superior progeny selected by any number ofselection methods, which include individual plant, half-sib progeny,full-sib progeny, and selfed progeny. The selected progeny arecross-pollinated with each other to form progeny for another population.This population is planted and again superior plants are selected tocross-pollinate with each other. Recurrent selection is a cyclicalprocess and therefore can be repeated as many times as desired. Theobjective of recurrent selection is to improve the traits of apopulation. The improved population can then be used as a source ofbreeding material to obtain new varieties for commercial or breedinguse, including the production of a synthetic variety. A syntheticvariety is the resultant progeny formed by the intercrossing of severalselected varieties.

Mass selection is a useful technique when used in conjunction withmolecular marker enhanced selection. In mass selection, seeds fromindividuals are selected based on phenotype or genotype. These selectedseeds are then bulked and used to grow the next generation. Bulkselection requires growing a population of plants in a bulk plot,allowing the plants to self-pollinate, harvesting the seed in bulk, andthen using a sample of the seed harvested in bulk to plant the nextgeneration. Also, instead of self-pollination, directed pollinationcould be used as part of the breeding program.

Mass and recurrent selections can be used to improve populations ofeither self- or cross-pollinating plants. A genetically variablepopulation of heterozygous individuals is either identified, or created,by intercrossing several different parents. The best plants are selectedbased on individual superiority, outstanding progeny, or excellentcombining ability. The selected plants are intercrossed to produce a newpopulation in which further cycles of selection are continued.

Mutation Breeding

Mutation breeding is another method of introducing new traits intoZantedeschia variety 110224-64275. Mutations that occur spontaneously orare artificially induced can be useful sources of variability for aplant breeder. The goal of artificial mutagenesis is to increase therate of mutation for a desired characteristic. Mutation rates can beincreased by many different means including temperature, long-term seedstorage, tissue culture conditions, radiation; such as X-rays, Gammarays (e.g., cobalt 60 or cesium 137), neutrons, (product of nuclearfission by uranium 235 in an atomic reactor), Beta radiation (emittedfrom radioisotopes such as phosphorus 32 or carbon 14), or ultravioletradiation (preferably from 2500 to 2900 nm), or chemical mutagens (suchas base analogues (5-bromo-uracil)), related compounds (8-ethoxycaffeine), antibiotics (streptonigrin), alkylating agents (sulfurmustards, nitrogen mustards, epoxides, ethylenamines, sulfates,sulfonates, sulfones, lactones), azide, hydroxylamine, nitrous acid, oracridines. Once a desired trait is observed through mutagenesis thetrait may then be incorporated into existing germplasm by traditionalbreeding techniques. Details of mutation breeding can be found in Fehr,“Principles of Cultivar Development,” Macmillan Publishing Company(1993). In addition, mutations created in other Zantedeschia plants maybe used to produce a backcross conversion of Zantedeschia variety110224-64275 that comprises such mutation.

Additional methods include, but are not limited to, expression vectorsintroduced into plant tissues using a direct gene transfer method, suchas microprojectile-mediated delivery, DNA injection, electroporation,and the like. More preferably, expression vectors are introduced intoplant tissues by using either microprojectile-mediated delivery with abiolistic device or by using Agrobacterium-mediated transformation.Transformant plants obtained with the protoplasm of the invention areintended to be within the scope of this invention.

Single-Gene Conversions

When the term Zantedeschia plant is used in the context of an embodimentof the present invention, this also includes any single gene conversionsof that variety. The term single gene converted plant as used hereinrefers to those Zantedeschia plants which are developed by a plantbreeding technique called backcrossing wherein essentially all of thedesired morphological and physiological characteristics of a variety arerecovered in addition to the single gene transferred into the varietyvia the backcrossing technique. Backcrossing methods can be used withone embodiment of the present invention to improve or introduce acharacteristic into the variety. The term “backcrossing” as used hereinrefers to the repeated crossing of a hybrid progeny back to therecurrent parent, i.e., backcrossing 1, 2, 3, 4, 5, 6, 7, 8, or moretimes to the recurrent parent. The parental Zantedeschia plant thatcontributes the gene for the desired characteristic is termed thenonrecurrent or donor parent. This terminology refers to the fact thatthe nonrecurrent parent is used one time in the backcross protocol andtherefore does not recur. The parental Zantedeschia plant to which thegene or genes from the nonrecurrent parent are transferred is known asthe recurrent parent as it is used for several rounds in thebackcrossing protocol (Poehlman & Sleper (1994); Fehr, Principles ofCultivar Development, pp. 261-286 (1987)). In a typical backcrossprotocol, the original variety of interest (recurrent parent) is crossedto a second variety (nonrecurrent parent) that carries the single geneof interest to be transferred. The resulting progeny from this cross arethen crossed again to the recurrent parent and the process is repeateduntil a Zantedeschia plant is obtained wherein essentially all of thedesired morphological and physiological characteristics of the recurrentparent are recovered in the converted plant, in addition to the singletransferred gene from the nonrecurrent parent.

The selection of a suitable recurrent parent is an important step for asuccessful backcrossing procedure. The goal of a backcross protocol isto alter or substitute a single trait or characteristic in the originalvariety. To accomplish this, a single gene of the recurrent variety ismodified or substituted with the desired gene from the nonrecurrentparent, while retaining essentially all of the rest of the desiredgenetic, and therefore the desired physiological and morphologicalconstitution of the original variety. The choice of the particularnonrecurrent parent will depend on the purpose of the backcross; one ofthe major purposes is to add some commercially important trait or traitsto the plant. The exact backcrossing protocol will depend on thecharacteristic or trait being altered to determine an appropriatetesting protocol. Although backcrossing methods are simplified when thecharacteristic being transferred is a dominant allele, a recessiveallele may also be transferred. In this instance it may be necessary tointroduce a test of the progeny to determine if the desiredcharacteristic has been successfully transferred.

Many single gene traits have been identified that are not regularlyselected for in the development of a new variety but that can beimproved by backcrossing techniques. Single gene traits may or may notbe transgenic. Examples of these traits include, but are not limited to,male sterility, waxy starch, herbicide resistance, resistance forbacterial, fungal, or viral disease, insect resistance, male fertility,and commercial characteristics such as flower color. These genes aregenerally inherited through the nucleus. Several of these single genetraits are described in U.S. Pat. Nos. 5,959,185, 5,973,234, 5,977,445,and 6,080,920, the disclosures of which are specifically herebyincorporated by reference.

Introduction of a New Trait or Locus into Zantedeschia Variety110224-64275

Zantedeschia variety 110224-64275 represents a new base genetic varietyinto which a new locus or trait may be introgressed. Directtransformation and backcrossing represent two important methods that canbe used to accomplish such an introgression. The term backcrossconversion and single locus conversion are used interchangeably todesignate the product of a backcrossing program.

Backcross Conversions of Zantedeschia Variety 110224-64275

A backcross conversion of Zantedeschia variety 110224-64275 occurs whenDNA sequences are introduced through backcrossing (Allard, “Principlesof Plant Breeding” (1960); and Fehr, “Breeding Methods for CultivarDevelopment,” 2^(nd) ed., Wilcox editor (1987)) with Zantedeschiavariety 110224-64275 utilized as the recurrent parent. Both naturallyoccurring and transgenic DNA sequences may be introduced throughbackcrossing techniques. A backcross conversion may produce a plant witha trait or locus conversion in at least two or more backcrosses,including at least 2 crosses, at least 3 crosses, at least 4 crosses, atleast 5 crosses, and the like. Molecular marker assisted breeding orselection may be utilized to reduce the number of backcrosses necessaryto achieve the backcross conversion. For example, see, Openshaw, S. J.,et al., Marker-assisted Selection in Backcross Breeding, ProceedingsSymposium of the Analysis of Molecular Data, Crop Science Society ofAmerica, Corvallis, Oreg. (August 1994), where it is demonstrated that abackcross conversion can be made in as few as two backcrosses.

The complexity of the backcross conversion method depends on the type oftrait being transferred (single genes or closely linked genes ascompared to unlinked genes), the level of expression of the trait, thetype of inheritance (cytoplasmic or nuclear), and the types of parentsincluded in the cross. It is understood by those of ordinary skill inthe art that for single gene traits that are relatively easy toclassify, the backcross method is effective and relatively easy tomanage. (See, Allard, “Principles of Plant Breeding” (1960); and Fehr,“Breeding Methods for Cultivar Development,” 2^(nd) ed., Wilcox editor(1987)). Desired traits that may be transferred through backcrossconversion include, but are not limited to, sterility (nuclear andcytoplasmic), fertility restoration, drought tolerance, nitrogenutilization, ornamental features, disease resistance (bacterial, fungal,or viral), insect resistance, and herbicide resistance. In addition, anintrogression site itself, such as an FRT site, Lox site, or other sitespecific integration site, may be inserted by backcrossing and utilizedfor direct insertion of one or more genes of interest into a specificplant variety. In some embodiments of the invention, the number of locithat may be backcrossed into Zantedeschia variety 110224-64275 is atleast 1, 2, 3, 4, or 5, and/or no more than 6, 5, 4, 3, or 2. A singlelocus may contain several transgenes, such as a transgene for diseaseresistance that, in the same expression vector, also contains atransgene for herbicide resistance. The gene for herbicide resistancemay be used as a selectable marker and/or as a phenotypic trait. Asingle locus conversion of site specific integration system allows forthe integration of multiple genes at the converted loci.

The backcross conversion may result from either the transfer of adominant allele or a recessive allele. Selection of progeny containingthe trait of interest is accomplished by direct selection for a traitassociated with a dominant allele. Transgenes or genes transferred viabackcrossing typically function as a dominant single gene trait and arerelatively easy to classify. Selection of progeny for a trait that istransferred via a recessive allele requires growing and selfing thefirst backcross generation to determine which plants carry the recessivealleles. Recessive traits may require additional progeny testing insuccessive backcross generations to determine the presence of the locusof interest. The last backcross generation is usually selfed to givepure breeding progeny for the gene(s) being transferred, although abackcross conversion with a stably introgressed trait may also bemaintained by further backcrossing to the recurrent parent withselection for the converted trait.

Along with selection for the trait of interest, progeny are selected forthe phenotype of the recurrent parent. The backcross is a form ofinbreeding, and the features of the recurrent parent are automaticallyrecovered after successive backcrosses. Poehlman, Breeding Field Crops,p. 204 (1987). Poehlman suggests from one to four or more backcrosses,but as noted above, the number of backcrosses necessary can be reducedwith the use of molecular markers. Other factors, such as a geneticallysimilar donor parent, may also reduce the number of backcrossesnecessary. As noted by Poehlman, backcrossing is easiest for simplyinherited, dominant, and easily recognized traits.

One process for adding or modifying a trait or locus in Zantedeschiavariety 110224-64275 comprises crossing Zantedeschia variety110224-64275 plants grown from Zantedeschia variety 110224-64275 seedwith plants of another Zantedeschia variety that comprise the desiredtrait or locus, selecting F₁ progeny plants that comprise the desiredtrait or locus to produce selected F₁ progeny plants, crossing theselected progeny plants with Zantedeschia variety 110224-64275 plants toproduce backcross progeny plants, selecting for backcross progeny plantsthat have the desired trait or locus and the morphologicalcharacteristics of Zantedeschia variety 110224-64275 to produce selectedbackcross progeny plants, and backcrossing to Zantedeschia variety110224-64275 three or more times in succession to produce selectedfourth or higher backcross progeny plants that comprise said trait orlocus. The modified Zantedeschia variety 110224-64275 may be furthercharacterized as having the physiological and morphologicalcharacteristics of Zantedeschia variety 110224-64275 listed in Table 1as determined at the 5% significance level when grown in the sameenvironmental conditions and/or may be characterized by percentsimilarity or identity to Zantedeschia variety 110224-64275 asdetermined by SSR markers. The above method may be utilized with fewerbackcrosses in appropriate situations, such as when the donor parent ishighly related or markers are used in the selection step. Desired traitsthat may be used include those nucleic acids known in the art, some ofwhich are listed herein, that will affect traits through nucleic acidexpression or inhibition. Desired loci include the introgression of FRT,Lox, and other sites for site specific integration, which may alsoaffect a desired trait if a functional nucleic acid is inserted at theintegration site.

In addition, the above process and other similar processes describedherein may be used to produce first generation progeny Zantedeschia seedby adding a step at the end of the process that comprises crossingZantedeschia variety 110224-64275 with the introgressed trait or locuswith a different Zantedeschia plant and harvesting the resultant firstgeneration progeny Zantedeschia seed.

Molecular Techniques Using Zantedeschia Variety 110224-64275

The advent of new molecular biological techniques has allowed theisolation and characterization of genetic elements with specificfunctions, such as encoding specific protein products. Scientists in thefield of plant biology developed a strong interest in engineering thegenome of plants to contain and express foreign genetic elements, oradditional, or modified versions of native or endogenous geneticelements in order to “alter” (the utilization of up-regulation,down-regulation, or gene silencing) the traits of a plant in a specificmanner. Any DNA sequences, whether from a different species or from thesame species, which are introduced into the genome using transformationor various breeding methods are referred to herein collectively as“transgenes.” In some embodiments of the invention, a transgenic variantof Zantedeschia variety 110224-64275 may contain at least one transgenebut could contain at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, and/or no morethan 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, or 2. Over the lastfifteen to twenty years several methods for producing transgenic plantshave been developed, and another embodiment of the present inventionalso relates to transgenic variants of Zantedeschia variety110224-64275.

Nucleic acids or polynucleotides refer to RNA or DNA that is linear orbranched, single or double stranded, or a hybrid thereof. The term alsoencompasses RNA/DNA hybrids. These terms also encompass untranslatedsequence located at both the 3′ and 5′ ends of the coding region of thegene: at least about 1000 nucleotides of sequence upstream from the 5′end of the coding region and at least about 200 nucleotides of sequencedownstream from the 3′ end of the coding region of the gene. Less commonbases, such as inosine, 5-methylcytosine, 6-methyladenine, hypoxanthineand others can also be used for antisense, dsRNA and ribozyme pairing.For example, polynucleotides that contain C-5 propyne analogues ofuridine and cytidine have been shown to bind RNA with high affinity andto be potent antisense inhibitors of gene expression. Othermodifications, such as modification to the phosphodiester backbone, orthe 2′-hydroxy in the ribose sugar group of the RNA can also be made.The antisense polynucleotides and ribozymes can consist entirely ofribonucleotides, or can contain mixed ribonucleotides anddeoxyribonucleotides. The polynucleotides of the invention may beproduced by any means, including genomic preparations, cDNApreparations, in-vitro synthesis, RT-PCR, and in-vitro or in-vivotranscription.

One embodiment of the invention is a process for producing Zantedeschiavariety 110224-64275 further comprising a desired trait, said processcomprising introducing a transgene that confers a desired trait to aZantedeschia plant of variety 110224-64275. Another embodiment is theproduct produced by this process. In one embodiment the desired traitmay be one or more of herbicide resistance, insect resistance, ordisease resistance. The specific gene may be any known in the art orlisted herein, including a polynucleotide conferring resistance toimidazolinone, dicamba, sulfonylurea, glyphosate, glufosinate, triazine,benzonitrile, cyclohexanedione, phenoxy proprionic acid, andL-phosphinothricin; a polynucleotide encoding a Bacillus thuringiensispolypeptide; or disease resistance to bacterial soft rot.

Numerous methods for plant transformation have been developed, includingbiological and physical plant transformation protocols. See, forexample, Miki et al., “Procedures for Introducing Foreign DNA intoPlants,” in Methods in Plant Molecular Biology and Biotechnology, andGlick and Thompson Eds., CRC Press, Inc., Boca Raton, pp. 67-88 (1993).In addition, expression vectors and in vitro culture methods for plantcell or tissue transformation and regeneration of plants are available.See, for example, Gruber, et al., “Vectors for Plant Transformation,” inMethods in Plant Molecular Biology and Biotechnology, Glick and ThompsonEds., CRC Press, Inc., Boca Raton, pp. 89-119 (1993).

A genetic trait which has been engineered into the genome of aparticular Zantedeschia plant may then be moved into the genome ofanother variety using traditional breeding techniques that are wellknown in the plant breeding arts. For example, a backcrossing approachis commonly used to move a transgene from a transformed Zantedeschiavariety into an already developed Zantedeschia variety, and theresulting backcross conversion plant would then comprise thetransgene(s).

Various genetic elements can be introduced into the plant genome usingtransformation. These elements include, but are not limited to, genes,coding sequences, inducible, constitutive and tissue specific promoters,enhancing sequences, and signal and targeting sequences. For example,see the traits, genes, and transformation methods listed in U.S. Pat.No. 6,118,055.

Breeding with Molecular Markers

Molecular markers, which includes markers identified through the use oftechniques such as Isozyme Electrophoresis, Restriction Fragment LengthPolymorphisms (RFLPs), Randomly Amplified Polymorphic DNAs (RAPDs),Arbitrarily Primed Polymerase Chain Reaction (AP-PCR), DNA AmplificationFingerprinting (DAF), Sequence Characterized Amplified Regions (SCARs),Amplified Fragment Length Polymorphisms (AFLPs), Simple Sequence Repeats(SSRs), and Single Nucleotide Polymorphisms (SNPs), may be used in plantbreeding methods utilizing Zantedeschia variety 110224-64275.

Isozyme electrophoresis and RFLPs have been widely used to determinegenetic composition. See R. L. Phillips and I. K. Vasil (ed.), DNA-basedmarkers in plants, Kluwer Academic Press Dordrecht, the Netherlands.

SSR technology is currently the most efficient and practical markertechnology. More marker loci can be routinely used, and more alleles permarker locus can be found, using SSRs in comparison to RFLPs. See forexample, Wills, D. M. et al., Theoretical and Applied Genetics 110:941-947 (2005). Single Nucleotide Polymorphisms may also be used toidentify the unique genetic composition of the invention and progenyvarieties retaining that unique genetic composition. Various molecularmarker techniques may be used in combination to enhance overallresolution.

One use of molecular markers is Quantitative Trait Loci (QTL) mapping.QTL mapping is the use of markers, which are known to be closely linkedto alleles that have measurable effects on a quantitative trait.Selection in the breeding process is based upon the accumulation ofmarkers linked to the positive effecting alleles and/or the eliminationof the markers linked to the negative effecting alleles from the plant'sgenome.

Molecular markers can also be used during the breeding process for theselection of qualitative traits. For example, markers closely linked toalleles or markers containing sequences within the actual alleles ofinterest can be used to select plants that contain the alleles ofinterest during a backcrossing breeding program. The markers can also beused to select for the genome of the recurrent parent and against thegenome of the donor parent. Using this procedure can minimize the amountof genome from the donor parent that remains in the selected plants. Itcan also be used to reduce the number of crosses back to the recurrentparent needed in a backcrossing program. The use of molecular markers inthe selection process is often called genetic marker enhanced selection.Molecular markers may also be used to identify and exclude certainsources of germplasm as parental varieties or ancestors of a plant byproviding a means of tracking genetic profiles through crosses.

Expression Vectors for Zantedeschia Transformation: Marker Genes

Plant transformation involves the construction of an expression vectorwhich will function in plant cells. Such a vector comprises DNAcomprising a gene under control of, or operatively linked to, aregulatory element (for example, a promoter). Expression vectors includeat least one genetic marker operably linked to a regulatory element (forexample, a promoter) that allows transformed cells containing the markerto be either recovered by negative selection, i.e., inhibiting growth ofcells that do not contain the selectable marker gene, or by positiveselection, i.e., screening for the product encoded by the geneticmarker. Many commonly used selectable marker genes for planttransformation are well-known in the transformation arts, and include,for example, genes that code for enzymes that metabolically detoxify aselective chemical agent which may be an antibiotic or an herbicide, orgenes that encode an altered target which is insensitive to theinhibitor. A few positive selection methods are also known in the art.

One commonly used selectable marker gene for plant transformation is theneomycin phosphotransferase II (nptII) gene which, when under thecontrol of plant regulatory signals, confers resistance to kanamycin.Fraley, et al., Proc. Natl. Acad. Sci. USA, 80:4803 (1983).

Another commonly used selectable marker gene is the hygromycinphosphotransferase gene which confers resistance to the antibiotichygromycin. Vanden Elzen, et al., Plant Mol. Biol., 5:299 (1985).

Additional selectable marker genes of bacterial origin that conferresistance to antibiotics include gentamycin acetyl transferase,streptomycin phosphotransferase and aminoglycoside-3′-adenyltransferase, the bleomycin resistance determinant (Hayford, et al.,Plant Physiol., 86:1216 (1988); Jones, et al., Mol. Gen. Genet., 210:86(1987); Svab, et al., Plant Mol. Biol., 14:197 (1990); Hille, et al.,Plant Mol. Biol., 7:171 (1986)). Other selectable marker genes conferresistance to herbicides such as glyphosate, glufosinate, or bromoxynil(Comai, et al., Nature, 317:741-744 (1985); Gordon-Kamm, et al., PlantCell, 2:603-618 (1990); Stalker, et al., Science, 242:419-423 (1988)).

Selectable marker genes for plant transformation not of bacterial origininclude, for example, mouse dihydrofolate reductase, plant5-enolpyruvylshikimate-3-phosphate synthase, and plant acetolactatesynthase (Eichholtz, et al., Somatic Cell Mol. Genet., 13:67 (1987);Shah, et al., Science, 233:478 (1986); Charest, et al., Plant Cell Rep.,8:643 (1990)).

Another class of marker genes for plant transformation requiresscreening of presumptively transformed plant cells, rather than directgenetic selection of transformed cells, for resistance to a toxicsubstance such as an antibiotic. These genes are particularly useful toquantify or visualize the spatial pattern of expression of a gene inspecific tissues and are frequently referred to as reporter genesbecause they can be fused to a gene or gene regulatory sequence for theinvestigation of gene expression. Commonly used genes for screeningpresumptively transformed cells include β-glucuronidase (GUS),β-galactosidase, luciferase, and chloramphenicol acetyltransferase(Jefferson, R. A., Plant Mol. Biol. Rep., 5:387 (1987); Teeri, et al.,EMBO J., 8:343 (1989); Koncz, et al., Proc. Natl. Acad. Sci. USA, 84:131(1987); DeBlock, et al., EMBO J., 3:1681 (1984)).

In-vivo methods for visualizing GUS activity that do not requiredestruction of plant tissue are available (Molecular Probes, Publication2908, IMAGENE GREEN, pp. 1-4 (1993); Naleway, et al., J. Cell Biol.,115:151a (1991)). However, these in-vivo methods for visualizing GUSactivity have not proven useful for recovery of transformed cellsbecause of low sensitivity, high fluorescent backgrounds, andlimitations associated with the use of luciferase genes as selectablemarkers.

More recently, a gene encoding Green Fluorescent Protein (GFP) has beenutilized as a marker for gene expression in prokaryotic and eukaryoticcells (Chalfie, et al., Science, 263:802 (1994)). GFP and mutants of GFPmay be used as screenable markers.

Expression Vectors for Zantedeschia Transformation: Promoters

Genes included in expression vectors must be driven by a nucleotidesequence comprising a regulatory element (for example, a promoter).Several types of promoters are well known in the transformation arts asare other regulatory elements that can be used alone or in combinationwith promoters.

As used herein, “promoter” includes reference to a region of DNAupstream from the start of transcription and involved in recognition andbinding of RNA polymerase and other proteins to initiate transcription.A “plant promoter” is a promoter capable of initiating transcription inplant cells. Examples of promoters under developmental control includepromoters that preferentially initiate transcription in certain tissues,such as leaves, roots, seeds, tubers, rhizome, xylem vessels, tracheids,or sclerenchyma. Such promoters are referred to as “tissue-preferred.”Promoters that initiate transcription only in a certain tissue arereferred to as “tissue-specific.” A “cell-type” specific promoterprimarily drives expression in certain cell types in one or more organs,for example, vascular cells in roots or leaves. An “inducible” promoteris a promoter which is under environmental control. Examples ofenvironmental conditions that may effect transcription by induciblepromoters include anaerobic conditions or the presence of light.Tissue-specific, tissue-preferred, cell-type specific, and induciblepromoters constitute the class of “non-constitutive” promoters. A“constitutive” promoter is a promoter that is active under mostenvironmental conditions.

A. Inducible Promoters: An inducible promoter is operably linked to agene for expression in Zantedeschia. Optionally, the inducible promoteris operably linked to a nucleotide sequence encoding a signal sequencewhich is operably linked to a gene for expression in Zantedeschia. Withan inducible promoter the rate of transcription increases in response toan inducing agent.

Any inducible promoter can be used in the instant invention. See, Ward,et al., Plant Mol. Biol., 22:361-366 (1993). Exemplary induciblepromoters include, but are not limited to, that from the ACEI systemwhich responds to copper (Mett, et al., Proc. Natl. Acad. Sci. USA,90:4567-4571 (1993)); In2 gene from maize which responds tobenzenesulfonamide herbicide safeners (Hershey, et al., Mol. GenGenetics, 227:229-237 (1991); Gatz, et al., Mol. Gen. Genetics,243:32-38 (1994)); or Tet repressor from Tn10 (Gatz, et al., Mol. Gen.Genetics, 227:229-237 (1991)). A particularly preferred induciblepromoter is a promoter that responds to an inducing agent to whichplants do not normally respond. An exemplary inducible promoter is theinducible promoter from a steroid hormone gene, the transcriptionalactivity of which is induced by a glucocorticosteroid hormone (Schena,et al., Proc. Natl. Acad. Sci. USA, 88:0421 (1991)).

B. Constitutive Promoters: A constitutive promoter is operably linked toa gene for expression in Zantedeschia or the constitutive promoter isoperably linked to a nucleotide sequence encoding a signal sequencewhich is operably linked to a gene for expression in Zantedeschia.

Many different constitutive promoters can be utilized in the instantinvention. Exemplary constitutive promoters include, but are not limitedto, the promoters from plant viruses such as the 35S promoter from CaMV(Odell, et al., Nature, 313:810-812 (1985)) and the promoters from suchgenes as rice actin (McElroy, et al., Plant Cell, 2:163-171 (1990));ubiquitin (Christensen, et al., Plant Mol. Biol., 12:619-632 (1989);Christensen, et al., Plant Mol. Biol., 18:675-689 (1992)); pEMU (Last,et al., Theor. Appl. Genet., 81:581-588 (1991)); MAS (Velten, et al.,EMBO J., 3:2723-2730 (1984)); and maize H3 histone (Lepetit, et al.,Mol. Gen. Genetics, 231:276-285 (1992); Atanassova, et al., PlantJournal, 2 (3):291-300 (1992)). The ALS promoter, Xbal/NcoI fragment 5′to the Brassica napus ALS3 structural gene (or a nucleotide sequencesimilarity to said Xbal/NcoI fragment), represents a particularly usefulconstitutive promoter. See, U.S. Pat. No. 5,659,026.

C. Tissue-Specific or Tissue-Preferred Promoters: A tissue-specificpromoter is operably linked to a gene for expression in Zantedeschia.Optionally, the tissue-specific promoter is operably linked to anucleotide sequence encoding a signal sequence which is operably linkedto a gene for expression in Zantedeschia. Plants transformed with a geneof interest operably linked to a tissue-specific promoter produce theprotein product of the transgene exclusively, or preferentially, in aspecific tissue.

Any tissue-specific or tissue-preferred promoter can be utilized in theinstant invention. Exemplary tissue-specific or tissue-preferredpromoters include, but are not limited to, a root-preferred promotersuch as that from the phaseolin gene (Murai, et al., Science, 23:476-482(1983); Sengupta-Gopalan, et al., Proc. Natl. Acad. Sci. USA,82:3320-3324 (1985)); a leaf-specific and light-induced promoter such asthat from cab or rubisco (Simpson, et al., EMBO J., 4 (11):2723-2729(1985); Timko, et al., Nature, 318:579-582 (1985)); an anther-specificpromoter such as that from LAT52 (Twell, et al., Mol. Gen. Genetics,217:240-245 (1989)); a pollen-specific promoter such as that from Zm13(Guerrero, et al., Mol. Gen. Genetics, 244:161-168 (1993)); or amicrospore-preferred promoter such as that from apg (Twell, et al., Sex.Plant Reprod., 6:217-224 (1993)).

Signal Sequences for Targeting Proteins to Subcellular Compartments

Transport of a protein produced by transgenes to a subcellularcompartment, such as the chloroplast, vacuole, peroxisome, glyoxysome,cell wall, or mitochondrion, or for secretion into the apoplast, isaccomplished by means of operably linking the nucleotide sequenceencoding a signal sequence to the 5′ and/or 3′ region of a gene encodingthe protein of interest. Targeting sequences at the 5′ and/or 3′ end ofthe structural gene may determine during protein synthesis andprocessing where the encoded protein is ultimately compartmentalized.

The presence of a signal sequence directs a polypeptide to either anintracellular organelle or subcellular compartment or for secretion tothe apoplast. Many signal sequences are known in the art. See, forexample, Becker, et al., Plant Mol. Biol., 20:49 (1992); Knox, C., etal., Plant Mol. Biol., 9:3-17 (1987); Lerner, et al., Plant Physiol.,91:124-129 (1989); Frontes, et al., Plant Cell, 3:483-496 (1991);Matsuoka, et al., Proc. Natl. Acad. Sci., 88:834 (1991); Gould, et al.,J. Cell. Biol., 108:1657 (1989); Creissen, et al., Plant J., 2:129(1991); Kalderon, et al., Cell, 39:499-509 (1984); Steifel, et al.,Plant Cell, 2:785-793 (1990).

Foreign Protein Genes and Ornamental Plant Genes: Transformation

With transgenic plants according to one embodiment of the presentinvention, a foreign protein can be produced in commercial quantities.Thus, techniques for the selection and propagation of transformedplants, which are well understood in the art, yield a plurality oftransgenic plants which are harvested in a conventional manner, and aforeign protein can then be extracted from a tissue of interest or fromtotal biomass. Protein extraction from plant biomass can be accomplishedby known methods which are discussed, for example, by Heney and Orr,Anal. Biochem., 114:92-6 (1981).

According to a preferred embodiment, the transgenic plant provided forcommercial production of foreign protein is a Zantedeschia plant. Inanother preferred embodiment, the biomass of interest is seed. For therelatively small number of transgenic plants that show higher levels ofexpression, a genetic map can be generated, primarily via conventionalRFLP, PCR, and SSR analysis, which identifies the approximatechromosomal location of the integrated DNA molecule. For exemplarymethodologies in this regard, see, Glick and Thompson, Methods in PlantMolecular Biology and Biotechnology, CRC Press, Inc., Boca Raton,269:284 (1993). Map information concerning chromosomal location isuseful for proprietary protection of a subject transgenic plant.

Wang, et al. discuss “Large Scale Identification, Mapping and Genotypingof Single-Nucleotide Polymorphisms in the Human Genome,” Science,280:1077-1082 (1998), and similar capabilities are becoming increasinglyavailable for the Zantedeschia genome. Map information concerningchromosomal location is useful for proprietary protection of a subjecttransgenic plant. If unauthorized propagation is undertaken and crossesmade with other germplasm, the map of the integration region can becompared to similar maps for suspect plants to determine if the latterhave a common parentage with the subject plant. Map comparisons wouldinvolve hybridizations, RFLP, PCR, SSR, and sequencing, all of which areconventional techniques. SNPs may also be used alone or in combinationwith other techniques.

Likewise, by means of one embodiment of the present invention, plantscan be genetically engineered to express various phenotypes ofcommercial interest. Through the transformation of Zantedeschia, theexpression of genes can be altered to enhance disease resistance, insectresistance, herbicide resistance, ornamental features, and other traits.Transformation can also be used to insert DNA sequences which control orhelp control male-sterility. DNA sequences native to Zantedeschia, aswell as non-native DNA sequences, can be transformed into Zantedeschiaand used to alter levels of native or non-native proteins. Variouspromoters, targeting sequences, enhancing sequences, and other DNAsequences can be inserted into the genome for the purpose of alteringthe expression of proteins. The interruption or suppression of theexpression of a gene at the level of transcription or translation (alsoknown as gene silencing or gene suppression) is desirable for severalaspects of genetic engineering in plants.

Many techniques for gene silencing are well-known to one of skill in theart, including, but not limited to, knock-outs (such as by insertion ofa transposable element such as Mu (Vicki Chandler, The Maize Handbook,Ch. 118 (Springer-Verlag 1994)) or other genetic elements such as a FRT,Lox, or other site specific integration sites; antisense technology(see, e.g., Sheehy, et al., PNAS USA, 85:8805-8809 (1988) and U.S. Pat.Nos. 5,107,065, 5,453,566, and 5,759,829); co-suppression (e.g., Taylor,Plant Cell, 9:1245 (1997); Jorgensen, Trends Biotech., 8 (12):340-344(1990); Flavell, PNAS USA, 91:3490-3496 (1994); Finnegan, et al.,Bio/Technology, 12:883-888 (1994); Neuhuber, et al., Mol. Gen. Genet.,244:230-241 (1994)); RNA interference (Napoli, et al., Plant Cell,2:279-289 (1990); U.S. Pat. No. 5,034,323; Sharp, Genes Dev., 13:139-141(1999); Zamore, et al., Cell, 101:25-33 (2000); Montgomery, et al., PNASUSA, 95:15502-15507 (1998)), virus-induced gene silencing (Burton, etal., Plant Cell, 12:691-705 (2000); Baulcombe, Curr. Op, Plant Bio.,2:109-113 (1999)); target-RNA-specific ribozymes (Haseloff, et al.,Nature, 334:585-591 (1988)); hairpin structures (Smith, et al., Nature,407:319-320 (2000); U.S. Pat. Nos. 6,423,885, 7,138,565, 6,753,139, and7,713,715); MicroRNA (Aukerman & Sakai, Plant Cell, 15:2730-2741(2003)); ribozymes (Steinecke, et al., EMBO J., 11:1525 (1992);Perriman, et al., Antisense Res. Dev., 3:253 (1993)); oligonucleotidemediated targeted modification (e.g., U.S. Pat. Nos. 6,528,700 and6,911,575); Zn-finger targeted molecules (e.g., U.S. Pat. Nos.7,151,201, 6,453,242, 6,785,613, 7,177,766 and 7,788,044); and othermethods or combinations of the above methods known to those of skill inthe art.

Methods for Zantedeschia Transformation

Numerous methods for plant transformation have been developed includingbiological and physical plant transformation protocols. See, forexample, Miki, et al., “Procedures for Introducing Foreign DNA intoPlants,” in Methods in Plant Molecular Biology and Biotechnology, Glickand Thompson Eds., CRC Press, Inc., Boca Raton, pp. 67-88 (1993). Inaddition, expression vectors and in-vitro culture methods for plant cellor tissue transformation and regeneration of plants are available. See,for example, Gruber, et al., “Vectors for Plant Transformation,” inMethods in Plant Molecular Biology and Biotechnology, Glick and ThompsonEds., CRC Press, Inc., Boca Raton, pp. 89-119 (1993).

A. Agrobacterium-mediated Transformation: One method for introducing anexpression vector into plants is based on the natural transformationsystem of Agrobacterium. See, for example, Horsch, et al., Science,227:1229 (1985). A. tumefaciens and A. rhizogenes are plant pathogenicsoil bacteria which genetically transform plant cells. The Ti and Riplasmids of A. tumefaciens and A. rhizogenes, respectively, carry genesresponsible for genetic transformation of the plant. See, for example,Kado, C. I., Crit. Rev. Plant Sci., 10:1 (1991). Descriptions ofAgrobacterium vector systems and methods for Agrobacterium-mediated genetransfer are provided by Gruber, et al., supra, Miki, et al., supra, andMoloney, et al., Plant Cell Reports, 8:238 (1989). See also, U.S. Pat.No. 5,563,055 (Townsend and Thomas), issued Oct. 8, 1996.

B. Direct Gene Transfer: Several methods of plant transformation,collectively referred to as direct gene transfer, have been developed asan alternative to Agrobacterium-mediated transformation. A generallyapplicable method of plant transformation is microprojectile-mediatedtransformation where DNA is carried on the surface of microprojectilesmeasuring 1 to 4 μm. The expression vector is introduced into planttissues with a biolistic device that accelerates the microprojectiles tospeeds of 300 to 600 m/s which is sufficient to penetrate plant cellwalls and membranes. Sanford, et al., Part. Sci. Technol., 5:27 (1987);Sanford, J. C., Trends Biotech., 6:299 (1988); Klein, et al., Bio/Tech.,6:559-563 (1988); Sanford, J. C., Physiol Plant, 7:206 (1990); Klein, etal., Biotechnology, 10:268 (1992). See also, U.S. Pat. No. 5,015,580(Christou, et al.), issued May 14, 1991 and U.S. Pat. No. 5,322,783.

Another method for physical delivery of DNA to plants is sonication oftarget cells. Zhang, et al., Bio/Technology, 9:996 (1991).Alternatively, liposome and spheroplast fusion have been used tointroduce expression vectors into plants. Deshayes, et al., EMBO J.,4:2731 (1985); Christou, et al., Proc Natl. Acad. Sci. USA, 84:3962(1987). Direct uptake of DNA into protoplasts using CaCl₂ precipitation,polyvinyl alcohol or poly-L-ornithine have also been reported. Hain, etal., Mol. Gen. Genet., 199:161 (1985) and Draper, et al., Plant CellPhysiol., 23:451 (1982). Electroporation of protoplasts and whole cellsand tissues has also been described (D'Halluin, et al., Plant Cell,4:1495-1505 (1992); and Spencer, et al., Plant Mol. Biol., 24:51-61(1994)).

Following transformation of Zantedeschia target tissues, expression ofthe above-described selectable marker genes allows for preferentialselection of transformed cells, tissues, and/or plants, usingregeneration and selection methods well-known in the art.

The foregoing methods for transformation would typically be used forproducing a transgenic variety. The transgenic variety could then becrossed with another (non-transformed or transformed) variety in orderto produce a new transgenic variety. Alternatively, a genetic trait thathas been engineered into a particular Zantedeschia plant using theforegoing transformation techniques could be moved into another lineusing traditional backcrossing techniques that are well-known in theplant breeding arts. For example, a backcrossing approach could be usedto move an engineered trait from a public, non-elite variety into anelite variety, or from a variety containing a foreign gene in its genomeinto a variety or varieties that do not contain that gene. As usedherein, “crossing” can refer to a simple x by y cross or the process ofbackcrossing depending on the context.

Likewise, by means of one embodiment of the present invention,commercially important genes can be expressed in transformed plants.More particularly, plants can be genetically engineered to expressvarious phenotypes of commercial interest. Exemplary genes implicated inthis regard include, but are not limited to, those categorized below:

1. Genes that Confer Resistance to Pests or Disease and that Encode:

A. Plant disease resistance genes. Plant defenses are often activated byspecific interaction between the product of a disease resistance gene(R) in the plant and the product of a corresponding avirulence (Avr)gene in the pathogen. A plant variety can be transformed with one ormore cloned resistance genes to engineer plants that are resistant tospecific pathogen strains. See, for example, Jones, et al., Science,266:789 (1994) (cloning of the tomato Cf-9 gene for resistance toCladosporium fulvum); Martin, et al., Science, 262:1432 (1993) (tomatoPto gene for resistance to Pseudomonas syringae pv. tomato encodes aprotein kinase); Mindrinos, et al., Cell, 78:1089 (1994) (ArabidopsisRSP2 gene for resistance to Pseudomonas syringae); McDowell & Woffenden,Trends Biotechnol., 21 (4):178-83 (2003); and Toyoda, et al., TransgenicRes., 11 (6):567-82 (2002).

B. A gene conferring resistance to a pest for spotted wilt or Dasheenmosaic virus.

C. A Bacillus thuringiensis protein, a derivative thereof or a syntheticpolypeptide modeled thereon. See, for example, Geiser, et al., Gene,48:109 (1986), who disclose the cloning and nucleotide sequence of a Btδ-endotoxin gene. Moreover, DNA molecules encoding δ-endotoxin genes canbe purchased from American Type Culture Collection, Manassas, Va., forexample, under ATCC Accession Nos. 40098, 67136, 31995, and 31998.

D. A vitamin-binding protein such as avidin. See, InternationalApplication No. PCT/US1993/006487, which teaches the use of avidin andavidin homologues as larvicides against insect pests.

E. An enzyme inhibitor, for example, a protease or proteinase inhibitoror an amylase inhibitor. See, for example, Abe, et al., J. Biol. Chem.,262:16793 (1987) (nucleotide sequence of rice cysteine proteinaseinhibitor); Huub, et al., Plant Molec. Biol., 21:985 (1993) (nucleotidesequence of cDNA encoding tobacco proteinase inhibitor I); Sumitani, etal., Biosci. Biotech. Biochem., 57:1243 (1993) (nucleotide sequence ofStreptomyces nitrosporeus α-amylase inhibitor); and U.S. Pat. No.5,494,813.

F. An insect-specific hormone or pheromone, such as an ecdysteroid orjuvenile hormone, a variant thereof, a mimetic based thereon, or anantagonist or agonist thereof. See, for example, the disclosure byHammock, et al., Nature, 344:458 (1990), of baculovirus expression ofcloned juvenile hormone esterase, an inactivator of juvenile hormone.

G. An insect-specific peptide or neuropeptide which, upon expression,disrupts the physiology of the affected pest. For example, see thedisclosures of Regan, J. Biol. Chem., 269:9 (1994) (expression cloningyields DNA coding for insect diuretic hormone receptor); Pratt, et al.,Biochem. Biophys. Res. Comm., 163:1243 (1989) (an allostatin isidentified in Diploptera puntata); Chattopadhyay, et al., CriticalReviews in Microbiology, 30 (1):33-54 (2004); Zjawiony, J. Nat. Prod.,67 (2):300-310 (2004); Carlini & Grossi-de-Sa, Toxicon, 40(11):1515-1539 (2002); Ussuf, et al., Curr Sci., 80 (7):847-853 (2001);Vasconcelos & Oliveira, Toxicon, 44 (4):385-403 (2004). See also, U.S.Pat. No. 5,266,317 which discloses genes encoding insect-specific,paralytic neurotoxins.

H. An insect-specific venom produced in nature by a snake, a wasp, etc.For example, see, Pang, et al., Gene, 116:165 (1992), for disclosure ofheterologous expression in plants of a gene coding for a scorpioninsectotoxic peptide.

I. An enzyme responsible for a hyperaccumulation of a monoterpene, asesquiterpene, a steroid, hydroxamic acid, a phenylpropanoid derivative,or another non-protein molecule with insecticidal activity.

J. An enzyme involved in the modification, including thepost-translational modification, of a biologically active molecule; forexample, a glycolytic enzyme, a proteolytic enzyme, a lipolytic enzyme,a nuclease, a cyclase, a transaminase, an esterase, a hydrolase, aphosphatase, a kinase, a phosphorylase, a polymerase, an elastase, achitinase, and a glucanase, whether natural or synthetic. See, U.S. Pat.No. 5,955,653 which discloses the nucleotide sequence of a callase gene.DNA molecules which contain chitinase-encoding sequences can beobtained, for example, from the ATCC under Accession Nos. 39637 and67152. See also, Kramer, et al., Insect Biochem. Molec. Biol., 23:691(1993), who teach the nucleotide sequence of a cDNA encoding tobaccohornworm chitinase, and Kawalleck, et al., Plant Molec. Biol., 21:673(1993), who provide the nucleotide sequence of the parsley ubi4-2polyubiquitin gene, U.S. Pat. Nos. 7,145,060, 7,087,810, and 6,563,020.

K. A molecule that stimulates signal transduction. For example, see thedisclosure by Botella, et al., Plant Molec. Biol., 24:757 (1994), ofnucleotide sequences for mung bean calmodulin cDNA clones, and Griess,et al., Plant Physiol., 104:1467 (1994), who provide the nucleotidesequence of a maize calmodulin cDNA clone.

L. A hydrophobic moment peptide. See, U.S. Pat. No. 5,580,852, whichdiscloses peptide derivatives of tachyplesin which inhibit fungal plantpathogens, and U.S. Pat. No. 5,607,914 which teaches syntheticantimicrobial peptides that confer disease resistance.

M. A membrane permease, a channel former or a channel blocker. Forexample, see the disclosure of Jaynes, et al., Plant Sci, 89:43 (1993),of heterologous expression of a cecropin-13 lytic peptide analog torender transgenic tobacco plants resistant to Pseudomonas solanacearum.

N. A viral-invasive protein or a complex toxin derived therefrom. Forexample, the accumulation of viral coat proteins in transformed plantcells imparts resistance to viral infection and/or disease developmenteffected by the virus from which the coat protein gene is derived, aswell as by related viruses. See, Beachy, et al., Ann. Rev. Phytopathol.,28:451 (1990). Coat protein-mediated resistance has been conferred upontransformed plants against alfalfa mosaic virus, cucumber mosaic virus,and tobacco mosaic virus.

O. An insect-specific antibody or an immunotoxin derived therefrom.Thus, an antibody targeted to a critical metabolic function in theinsect gut would inactivate an affected enzyme, killing the insect.

P. A virus-specific antibody. See, for example, Tavladoraki, et al.,Nature, 366:469 (1993), who show that transgenic plants expressingrecombinant antibody genes are protected from virus attack.

Q. A developmental-arrestive protein produced in nature by a pathogen ora parasite. Thus, fungal endo-α-1,4-D-polygalacturonases facilitatefungal colonization and plant nutrient release by solubilizing plantcell wall homo-α-1,4-D-galacturonase. See, Lamb, et al., Bio/Technology,10:1436 (1992). The cloning and characterization of a gene which encodesa bean endopolygalacturonase-inhibiting protein is described by Toubart,et al., Plant J., 2:367 (1992).

Q. A developmental-arrestive protein produced in nature by a plant. Forexample, Logemann, et al., Bio/Technology, 10:305 (1992), have shownthat transgenic plants expressing the barley ribosome-inactivating genehave an increased resistance to fungal disease.

R. Genes involved in the Systemic Acquired Resistance (SAR) Responseand/or the pathogenesis-related genes. Briggs, S., Current Biology, 5(2) (1995); Pieterse & Van Loon, Curr. Opin. Plant Bio., 7 (4):456-64(2004); and Somssich, Cell, 113 (7):815-6 (2003).

S. Antifungal genes. See, Cornelissen and Melchers, Plant Physiol.,101:709-712 (1993); Parijs, et al., Planta, 183:258-264 (1991); andBushnell, et al., Can. J. of Plant Path., 20 (2):137-149 (1998). Seealso, U.S. Pat. No. 6,875,907.

T. Detoxification genes, such as for fumonisin, beauvericin,moniliformin, and zearalenone and their structurally-relatedderivatives. See, U.S. Pat. No. 5,792,931.

U. Cystatin and cysteine proteinase inhibitors. See, U.S. Pat. No.7,205,453.

V. Genes that confer resistance to Phytophthora Root Rot, such as theRps 1, Rps 1-a, Rps 1-b, Rps 1-c, Rps 1-d, Rps 1-e, Rps 1-k, Rps 2, Rps3-a, Rps 3-b, Rps 3-c, Rps 4, Rps 5, Rps 6, Rps 7, and other Rps genes.

W. Genes that confer resistance to Pythium root rot and crown rot. Seefor example, Tanyolac, Bahattin, et al., African J. of Biotech. 9(19):2727-2730 (2011) and Nzungize, J., et al., African J. of Biotech. 5(3):193-200 (2011).

Any of the above-listed disease or pest resistance genes (A-W) can beintroduced into Zantedeschia variety 110224-64275 through a variety ofmeans including, but not limited to, transformation and crossing.

2. Genes that Confer Resistance to an Herbicide, for Example:

A. An herbicide that inhibits the growing point or meristem, such as animidazolinone or a sulfonylurea. Exemplary genes in this category codefor mutant ALS and AHAS enzyme as described, for example, by Lee, etal., EMBO J., 7:1241 (1988) and Mild, et al., Theor. Appl. Genet.,80:449 (1990), respectively.

B. Glyphosate (resistance conferred by mutant5-enolpyruvlshikimate-3-phosphate synthase (EPSPS) and aroA genes,respectively) and other phosphono compounds, such as glufosinate(phosphinothricin acetyl transferase (PAT) and Streptomyceshygroscopicus PAT bar genes), pyridinoxy or phenoxy proprionic acids,and cyclohexanediones (ACCase inhibitor-encoding genes). See, forexample, U.S. Pat. No. 4,940,835 which discloses the nucleotide sequenceof a form of EPSPS which can confer glyphosate resistance. U.S. Pat. No.5,627,061 which describes genes encoding EPSPS enzymes. See also, U.S.Pat. Nos. 6,566,587, 6,338,961, 6,248,876, 6,040,497, 5,804,425,5,633,435, 5,145,783, 4,971,908, 5,312,910, 5,188,642, 4,940,835,5,866,775, 6,225,114, 6,130,366, 5,310,667, 4,535,060, 4,769,061,5,633,448, 5,510,471, 6,803,501, RE 36,449, RE 37,287, and 5,491,288,which are incorporated herein by reference for this purpose. Glyphosateresistance is also imparted to plants that express a gene that encodes aglyphosate oxido-reductase enzyme, as described more fully in U.S. Pat.Nos. 5,776,760 and 5,463,175, which are incorporated herein by referencefor this purpose. In addition, glyphosate resistance can be imparted toplants by the over expression of genes encoding glyphosateN-acetyltransferase. See, for example, U.S. Pat. No. 7,462,481. A DNAmolecule encoding a mutant aroA gene can be obtained under ATCCAccession No. 39256, and the nucleotide sequence of the mutant gene isdisclosed in U.S. Pat. No. 4,769,061. European Patent Appl. No. 0333033and U.S. Pat. No. 4,975,374 disclose nucleotide sequences of glutaminesynthetase genes which confer resistance to herbicides such asL-phosphinothricin. The nucleotide sequence of a PAT gene is provided inEuropean Patent No. 0242246 to Leemans, et al. DeGreef, et al.,Bio/Technology, 7:61 (1989) describe the production of transgenic plantsthat express chimeric bar genes coding for phosphinothricin acetyltransferase activity. Exemplary of genes conferring resistance tophenoxy proprionic acids and cyclohexones, such as sethoxydim andhaloxyfop are the Acc1-S1, Acc1-S2, and Acc2-S3 genes described byMarshall, et al., Theor. Appl. Genet., 83:435 (1992).

C. An herbicide that inhibits photosynthesis, such as a triazine (psbAand gs+ genes) and a benzonitrile (nitrilase gene). Przibila, et al.,Plant Cell, 3:169 (1991), describe the transformation of Chlamydomonaswith plasmids encoding mutant psbA genes. Nucleotide sequences fornitrilase genes are disclosed in U.S. Pat. No. 4,810,648 to Stalker andDNA molecules containing these genes are available under ATCC AccessionNos. 53435, 67441, and 67442. Cloning and expression of DNA coding for aglutathione S-transferase is described by Hayes, et al., Biochem. J.,285:173 (1992).

D. Acetohydroxy acid synthase, which has been found to make plants thatexpress this enzyme resistant to multiple types of herbicides, has beenintroduced into a variety of plants. See, Hattori, et al., Mol. Gen.Genet., 246:419 (1995). Other genes that confer tolerance to herbicidesinclude a gene encoding a chimeric protein of rat cytochrome P4507A1 andyeast NADPH-cytochrome P450 oxidoreductase (Shiota, et al., PlantPhysiol., 106:17 (1994)); genes for glutathione reductase and superoxidedismutase (Aono, et al., Plant Cell Physiol., 36:1687 (1995)); and genesfor various phosphotransferases (Datta, et al., Plant Mol. Biol., 20:619(1992)).

E. Protoporphyrinogen oxidase (protox) is necessary for the productionof chlorophyll, which is necessary for all plant survival. The protoxenzyme serves as the target for a variety of herbicidal compounds. Theseherbicides also inhibit growth of all the different species of plantspresent, causing their total destruction. The development of plantscontaining altered protox activity which are resistant to theseherbicides are described in U.S. Pat. Nos. 6,288,306, 6,282,837,5,767,373, and 6,084,155.

Any of the above listed herbicide genes (A-E) can be introduced into theclaimed Zantedeschia variety through a variety of means including butnot limited to transformation and crossing.

3. Genes that Control Male Sterility:

There are several methods of conferring genetic male sterilityavailable, such as multiple mutant genes at separate locations withinthe genome that confer male sterility, as disclosed in U.S. Pat. Nos.4,654,465 and 4,727,219 to Brar, et al., and chromosomal translocationsas described by Patterson in U.S. Pat. Nos. 3,861,709 and 3,710,511. Inaddition to these methods, Albertsen, et al., U.S. Pat. No. 5,432,068,describes a system of nuclear male sterility which includes: identifyinga gene which is critical to male fertility; silencing this native genewhich is critical to male fertility; removing the native promoter fromthe essential male fertility gene and replacing it with an induciblepromoter; inserting this genetically engineered gene back into theplant; and thus creating a plant that is male sterile because theinducible promoter is not “on” resulting in the male fertility gene notbeing transcribed. Fertility is restored by inducing, or turning “on,”the promoter, which in turn allows the gene that confers male fertilityto be transcribed.

A. Introduction of a deacetylase gene under the control of atapetum-specific promoter and with the application of the chemicalN-Ac-PPT. See, U.S. Pat. No. 6,384,304.

B. Introduction of various stamen-specific promoters. See, U.S. Pat.Nos. 5,639,948 and 5,589,610.

C. Introduction of the barnase and the barstar genes. See, Paul, et al.,Plant Mol. Biol., 19:611-622 (1992).

For additional examples of nuclear male and female sterility systems andgenes, see also, U.S. Pat. Nos. 5,859,341, 6,297,426, 5,478,369,5,824,524, 5,850,014, and 6,265,640, all of which are herebyincorporated by reference.

4. Genes that Create a Site for Site Specific DNA Integration:

This includes the introduction of FRT sites that may be used in theFLP/FRT system and/or Lox sites that may be used in the Cre/Loxp system.See, for example, Lyznik, et al., Site-Specific Recombination forGenetic Engineering in Plants, Plant Cell Rep, 21:925-932 (2003) andU.S. Pat. No. 6,187,994, which are hereby incorporated by reference.Other systems that may be used include the Gin recombinase of phage Mu(Maeser, et al. (1991); the Pin recombinase of E. coli (Enomoto, et al.(1983)); and the R/RS system of the pSRi plasmid (Araki, et al. (1992)).

5. Genes that Affect Abiotic Stress Resistance:

Genes that affect abiotic stress resistance (including but not limitedto flowering, seed development, enhancement of nitrogen utilizationefficiency, altered nitrogen responsiveness, drought resistance ortolerance, cold resistance or tolerance, and salt resistance ortolerance). For example, see U.S. Pat. No. 6,653,535 where water useefficiency is altered through alteration of malate; U.S. Pat. Nos.5,892,009, 5,965,705, 5,929,305, 5,891,859, 6,417,428, 6,664,446,6,706,866, 6,717,034, 6,801,104, 6,946,586, 7,238,860, 7,635,800,7,135,616, 7,193,129, and 7,601,893; and International Publ. Nos. WO2001/026459, WO 2001/035725, WO 2001/035727, WO 2001/036444, WO2001/036597, WO 2001/036598, WO 2002/015675, and WO 2002/077185,describing genes, including CBF genes and transcription factorseffective in mitigating the negative effects of freezing, high salinity,and drought on plants, as well as conferring other positive effects onplant phenotype; U.S. Publ. No. 2004/0148654, where abscisic acid isaltered in plants resulting in improved plant phenotype, such asincreased tolerance to abiotic stress; U.S. Pat. Nos. 6,992,237,6,429,003, 7,049,115, and 7,262,038, where cytokinin expression ismodified resulting in plants with increased stress tolerance, such asdrought tolerance. See also, WO 02/02776, WO 2003/052063, JP 2002281975,U.S. Pat. No. 6,084,153, WO 01/64898, and U.S. Pat. Nos. 6,177,275 and6,107,547 (enhancement of nitrogen utilization and altered nitrogenresponsiveness). For ethylene alteration, see, U.S. Publ. Nos.2004/0128719, 2003/0166197, and U.S. application Ser. No. 09/856,834.For plant transcription factors or transcriptional regulators of abioticstress, see, e.g., U.S. Publ. Nos. 2004/0098764 or 2004/0078852.

Other genes and transcription factors that affect plant growth andcommercial traits, such as flowering, plant growth, and/or plantstructure, can be introduced or introgressed into plants. See forexample, U.S. Pat. Nos. 6,140,085, and 6,265,637 (CO); U.S. Pat. No.6,670,526 (ESD4); U.S. Pat. Nos. 6,573,430 and 7,157,279 (TFL); U.S.Pat. No. 6,713,663 (FT); U.S. Pat. Nos. 6,794,560, 6,307,126 (GAI); U.S.Pat. No. 7,045,682 (VRN1); U.S. Pat. Nos. 6,949,694 and 7,253,274(VRN2); U.S. Pat. No. 6,887,708 (GI); U.S. Pat. No. 7,320,158 (FRI);U.S. Pat. No. 6,307,126 (GAI); U.S. Pat. Nos. 6,762,348 and 7,268,272(D8 and Rht); and U.S. Pat. Nos. 7,345,217, 7,511,190, 7,659,446, and7,825,296 (transcription factors).

Genetic Marker Profile Through SSR and First Generation Progeny

In addition to phenotypic observations, a plant can also be identifiedby its genotype. The genotype of a plant can be characterized through agenetic marker profile which can identify plants of the same variety, ora related variety, or be used to determine or validate a pedigree.Genetic marker profiles can be obtained by techniques such asRestriction Fragment Length Polymorphisms (RFLPs), Randomly AmplifiedPolymorphic DNAs (RAPDs), Arbitrarily Primed Polymerase Chain Reaction(AP-PCR), DNA Amplification Fingerprinting (DAF), Sequence CharacterizedAmplified Regions (SCARs), Amplified Fragment Length Polymorphisms(AFLPs), Simple Sequence Repeats (SSRs) (which are also referred to asMicrosatellites), and Single Nucleotide Polymorphisms (SNPs). Forexample, see, Berry, et al., “Assessing Probability of Ancestry UsingSimple Sequence Repeat Profiles: Applications to Maize Inbred Lines andSoybean Varieties,” Genetics, 165:331-342 (2003), each of which areincorporated by reference herein in their entirety.

Particular markers used for these purposes are not limited to anyparticular set of markers, but are envisioned to include any type ofmarker and marker profile which provides a means of distinguishingvarieties. One method of comparison is to use only homozygous loci forZantedeschia variety 110224-64275.

Primers and PCR protocols for assaying these and other markers arewell-known. In addition to being used for identification of Zantedeschiavariety 110224-64275, and plant parts and plant cells of Zantedeschiavariety 110224-64275, the genetic profile may be used to identify aZantedeschia plant produced through the use of Zantedeschia variety110224-64275 or to verify a pedigree for progeny plants produced throughthe use of Zantedeschia variety 110224-64275. The genetic marker profileis also useful in breeding and developing backcross conversions.

One embodiment of the present invention comprises a Zantedeschia plantcharacterized by molecular and physiological data obtained from therepresentative sample of said variety deposited with the American TypeCulture Collection (ATCC) or with the National Collections ofIndustrial, Food and Marine Bacteria (NCIMB). Further provided byanother embodiment of the invention is a Zantedeschia plant formed bythe combination of the disclosed Zantedeschia plant or plant cell withanother Zantedeschia plant or cell and comprising the homozygous allelesof the variety. “Cell” as used herein includes a plant cell, whetherisolated, in tissue culture or incorporated in a plant or plant part

Means of performing genetic marker profiles using SSR polymorphisms arewell-known in the art. SSRs are genetic markers based on polymorphismsin repeated nucleotide sequences, such as microsatellites. A markersystem based on SSRs can be highly informative in linkage analysisrelative to other marker systems in that multiple alleles may be present(“linkage” refers to a phenomenon wherein alleles on the same chromosometend to segregate together more often than expected by chance if theirtransmission was independent). Another advantage of this type of markeris that, through use of flanking primers, detection of SSRs can beachieved, for example, by the polymerase chain reaction (PCR), therebyeliminating the need for labor-intensive Southern hybridization. The PCRdetection is done by use of two oligonucleotide primers flanking thepolymorphic segment of repetitive DNA. Repeated cycles of heatdenaturation of the DNA followed by annealing of the primers to theircomplementary sequences at low temperatures, and extension of theannealed primers with DNA polymerase, comprise the major part of themethodology.

Following amplification, markers can be scored by electrophoresis of theamplification products. Scoring of marker genotype is based on the sizeof the amplified fragment, which may be measured by the number of basepairs of the fragment. While variation in the primer used or inlaboratory procedures can affect the reported fragment size, relativevalues should remain constant regardless of the specific primer orlaboratory used. When comparing varieties it is preferable if all SSRprofiles are performed in the same lab. Primers used are publiclyavailable and well-known.

The SSR profile of a Zantedeschia plant 110224-64275 can be used toidentify plants comprising Zantedeschia variety 110224-64275 as aparent, since such plants will comprise the same homozygous alleles asZantedeschia variety 110224-64275. Because the Zantedeschia variety isessentially homozygous at all relevant loci, most loci should have onlyone type of allele present. In contrast, a genetic marker profile of anF₁ progeny should be the sum of those parents, e.g., if one parent washomozygous for allele x at a particular locus, and the other parenthomozygous for allele y at that locus, then the F₁ progeny will be xy(heterozygous) at that locus. Subsequent generations of progeny producedby selection and breeding are expected to be of genotype x (homozygous),y (homozygous), or xy (heterozygous) for that locus position. When theF₁ plant is selfed or sibbed for successive filial generations, thelocus should be either x or y for that position.

In addition, plants and plant parts substantially benefiting from theuse of Zantedeschia variety 110224-64275 in their development, such asZantedeschia variety 110224-64275 comprising a backcross conversion,transgene, or genetic sterility factor, may be identified by having amolecular marker profile with a high percent identity to Zantedeschiavariety 110224-64275. Such a percent identity might be 95%, 96%, 97%,98%, 99%, 99.5%, or 99.9% identical to Zantedeschia variety110224-64275. Percent identity refers to the comparison of thehomozygous alleles of two Zantedeschia varieties. Percent identity orpercent similarity is determined by comparing a statisticallysignificant number of the homozygous alleles of two developed varieties.For example, a percent identity of 90% between Zantedeschia variety 1and Zantedeschia variety 2 means that the two varieties have the sameallele at 90% of their loci.

The SSR profile of Zantedeschia variety 110224-64275 can also be used toidentify essentially derived varieties and other progeny varietiesdeveloped from the use of Zantedeschia variety 110224-64275, as well ascells and other plant parts thereof. Such plants may be developed usingthe markers, for example, identified in U.S. Pat. Nos. 6,162,967, and7,288,386. Progeny plants and plant parts produced using Zantedeschiavariety 110224-64275 may be identified by having a molecular markerprofile of at least 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%,75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%,89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 99.5% geneticcontribution from Zantedeschia variety, as measured by either percentidentity or percent similarity. Such progeny may be furthercharacterized as being within a pedigree distance of Zantedeschiavariety 110224-64275, such as within 1, 2, 3, 4, or 5 or lesscross-pollinations to a Zantedeschia plant other than Zantedeschia110224-64275 or a plant that has Zantedeschia variety 110224-64275 as aprogenitor. Unique molecular profiles may be identified with othermolecular tools such as SNPs and RFLPs.

While determining the SSR genetic marker profile of the plants describedsupra, several unique SSR profiles may also be identified which did notappear in either parent of such plant. Such unique SSR profiles mayarise during the breeding process from recombination or mutation. Acombination of several unique alleles provides a means of identifying aplant variety, an F₁ progeny produced from such variety, and progenyproduced from such variety.

Tissue Culture

Further reproduction of the variety can occur by tissue culture andregeneration. Tissue culture of various tissues of ornamental plants andZantedeschia and regeneration of plants therefrom is well-known andwidely published. For example, reference may be had to do Valla Rego,Luciana et al., Crop Breeding and Applied Technology. 1 (3): 283-300(2001); Komatsuda, T., et al., Crop Sci., 31:333-337 (1991); Stephens,P. A., et al., Theor. Appl. Genet., 82:633-635 (1991); Komatsuda, T., etal., Plant Cell, Tissue and Organ Culture, 28:103-113 (1992); Dhir, S.,et al., Plant Cell Reports, 11:285-289 (1992); Pandey, P., et al., JapanJ. Breed., 42:1-5 (1992); and Shetty, K., et al., Plant Science,81:245-251 (1992). Thus, another aspect of this invention is to providecells which upon growth and differentiation produce Zantedeschia plantshaving the physiological and morphological characteristics ofZantedeschia variety 110224-64275.

Regeneration refers to the development of a plant from tissue culture.The term “tissue culture” indicates a composition comprising isolatedcells of the same or a different type or a collection of such cellsorganized into parts of a plant. Exemplary types of tissue cultures areprotoplasts, calli, plant clumps, and plant cells that can generatetissue culture that are intact in plants or parts of plants, such aspollen, ovules, embryos, protoplasts, meristematic cells, callus,pollen, leaves, ovules, anthers, cotyledons, hypocotyl, pistils, roots,root tips, spathe, spadix, seeds, petiole, rhizome, pods, berries, bulb,tuber, shoot, or stems, and the like. Means for preparing andmaintaining plant tissue culture are well-known in the art.

While a number of exemplary aspects and embodiments have been discussedabove, those of skill in the art will recognize certain modifications,permutations, additions and sub-combinations thereof. It is thereforeintended that the following appended claims and claims hereafterintroduced are interpreted to include all such modifications,permutations, additions, and sub-combinations as are within their truespirit and scope.

The present invention may be embodied in other specific forms withoutdeparting from its spirit or essential characteristics. The describedembodiments are to be considered in all respects only as illustrativeand not restrictive. The scope of the invention is, therefore, indicatedby the appended claims rather than by the foregoing description. Allchanges which come within the meaning and range of equivalency of theclaims are to be embraced within their scope.

The present invention, in various embodiments, include components,methods, processes, systems and/or apparatus substantially as depictedand described herein, including various embodiments, subcombinations,and subsets thereof. Those of skill in the art will understand how tomake and use the present invention after understanding the presentdisclosure.

The foregoing discussion of the invention has been presented forpurposes of illustration and description. The foregoing is not intendedto limit the invention to the form or forms disclosed herein. In theforegoing Detailed Description for example, various features of theinvention are grouped together in one or more embodiments for thepurpose of streamlining the disclosure. This method of disclosure is notto be interpreted as reflecting an intention that the claimed inventionrequires more features than are expressly recited in each claim. Rather,as the following claims reflect, inventive aspects lie in less than allfeatures of a single foregoing disclosed embodiment. Thus, the followingclaims are hereby incorporated into this Detailed Description, with eachclaim standing on its own as a separate preferred embodiment of theinvention.

Moreover, though the description of the invention has includeddescription of one or more embodiments and certain variations andmodifications, other variations and modifications are within the scopeof the invention (e.g., as may be within the skill and knowledge ofthose in the art, after understanding the present disclosure). It isintended to obtain rights which include alternative embodiments to theextent permitted, including alternate, interchangeable and/or equivalentstructures, functions, ranges or acts to those claimed, whether or notsuch alternate, interchangeable and/or equivalent structures, functions,ranges or acts are disclosed herein, and without intending to publiclydedicate any patentable subject matter.

The use of the terms “a,” “an,” and “the,” and similar referents in thecontext of describing the invention (especially in the context of thefollowing claims) are to be construed to cover both the singular and theplural, unless otherwise indicated herein or clearly contradicted bycontext. The terms “comprising,” “having,” “including,” and “containing”are to be construed as open-ended terms (i.e., meaning “including, butnot limited to,”) unless otherwise noted. Recitation of ranges of valuesherein are merely intended to serve as a shorthand method of referringindividually to each separate value falling within the range, unlessotherwise indicated herein, and each separate value is incorporated intothe specification as if it were individually recited herein. Forexample, if the range 10-15 is disclosed, then 11, 12, 13, and 14 arealso disclosed. All methods described herein can be performed in anysuitable order unless otherwise indicated herein or otherwise clearlycontradicted by context. The use of any and all examples, or exemplarylanguage (e.g., “such as”) provided herein, is intended merely to betterilluminate the invention and does not pose a limitation on the scope ofthe invention unless otherwise claimed. No language in the specificationshould be construed as indicating any non-claimed element as essentialto the practice of the invention.

DEPOSIT INFORMATION

A deposit of the Golden State Bulb Growers, Inc. proprietaryZantedeschia variety 110224-64275 disclosed above and recited in theappended claims has been made with the National Collections ofIndustrial, Food and Marine Bacteria (NCIMB), Ferguson Building,Craibstone Estate, Bucksburn, Aberdeen, Scotland, AB21 9YA, UnitedKingdom. The date of deposit was Oct. 18, 2012. The deposit of 2,500seeds was taken from the same deposit maintained by Golden State BulbGrowers, Inc. since prior to the filing date of this application. Allrestrictions will be removed upon granting of a patent, and the depositis intended to meet all of the requirements of 37 C.F.R. §§1.801-1.809.The NCIMB No. is 42073. The deposit will be maintained in the depositoryfor a period of thirty years, or five years after the last request, orfor the enforceable life of the patent, whichever is longer, and will bereplaced as necessary during the period.

What is claimed is:
 1. A seed of Zantedeschia variety 110224-64275,wherein a representative sample of seed of said variety is depositedunder NCIMB No.
 42073. 2. A Zantedeschia plant, or a part thereof,produced by growing the seed of claim
 1. 3. A tissue culture producedfrom protoplasts or cells from the plant of claim 2, wherein said cellsor protoplasts are produced from a plant part selected from the groupconsisting of pollen, ovules, embryos, protoplasts, meristematic cells,callus, pollen, leaves, ovules, anthers, cotyledons, hypocotyl, pistils,roots, root tips, spathe, spadix, seeds, petiole, rhizome, pods,berries, bulb, tuber, shoot, and stems.
 4. A Zantedeschia plant ofvariety 110224-64275 regenerated from the tissue culture of claim
 3. 5.A method for producing a Zantedeschia seed, said method comprisingcrossing two Zantedeschia plants and harvesting the resultantZantedeschia seed, wherein at least one Zantedeschia plant is theZantedeschia plant of claim
 2. 6. A method of producing an herbicideresistant Zantedeschia plant, wherein said method comprises introducinga gene conferring herbicide resistance into the plant of claim
 2. 7. Anherbicide resistant Zantedeschia plant produced by the method of claim6.
 8. A method of producing a pest or insect resistant Zantedeschiaplant, wherein said method comprises introducing a gene conferring pestor insect resistance into the Zantedeschia plant of claim
 2. 9. A pestor insect resistant Zantedeschia plant produced by the method of claim8.
 10. The Zantedeschia plant of claim 9, wherein the gene encodes aBacillus thuringiensis (Bt) endotoxin.
 11. A method of producing adisease resistant Zantedeschia plant, wherein said method comprisesintroducing a gene which confers disease resistance into theZantedeschia plant of claim
 2. 12. A disease resistant Zantedeschiaplant produced by the method of claim claim
 11. 13. A plant ofZantedeschia variety 110224-64275, wherein a representative sample ofseed of said variety is deposited under NCIMB No. 42073.